Project Summary The molecular mechanism that underlies evolution to post-traumatic epilepsy (PTE) in individuals with traumatic brain injury (TBI) is unknown. It has been suggested that the development of PTE following TBI is the combined result of kindling epileptogenesis during the early stages of recovery and triggering by residual irritation from damaged neural tissue. Glutamate, the major excitatory neurotransmitter (NT) in the central nervous system, must be tightly regulated to allow for proper neuronal signaling without creating excessive excitation. TBI produces an increase in glutamate release into the extracellular space and a concurrent decrease in the ability of excitatory amino acid transporters to reuptake glutamate that may lead to its accumulation, propagating secondary damage cascades and producing aberrant neuronal signaling. We suggest that the enhanced hippocampal glutamate levels seen post-TBI are in part due to changes of the neurosecretory machinery and SNARE regulators and/or re-sequestration process. Left unchecked, these changes become drivers of post-TBI epileptogenesis leading to PTE. The hypothesis that pathologic adaptation of inherent hippocampal glutamatergic synaptic plasticity promotes post-TBI epileptogenesis will be tested through three specific aims in animals (and appropriate controls) that have been administered a lateral fluid percussion injury (LFPI) and then monitored by telemetry until electroencephalographic (EEG) evidence of epileptiform activity: (1) Measure of tonic glutamate levels and evoked glutamateamate release in the hippocampal formation of epileptic and non-epileptic animals after moderate LFPI through use of a novel microarray electrode system. In tissue from hippocampal formation of these same animals we will use Western blotting techniques to quantify (2) components of the neurotransmitter release process and (3) certain glutamate transporters. Investigation of this putative relationship between aberrant glutamate neuronal exocytosis/resequestration and PTE will provide insight into potential mechanisms by which glutamate neurotransmission is regulated and into the molecular events of epileptogenesis as well as provide potential foci for therapeutic intervention. The long-term goal of this project is to restore to normal aberrant mechanisms associated with TBI-associated epileptogenesis. There are indications from animal studies that several newer FDA-approved and actively used antiepileptic drugs, e.g. levetiracetam and lacosamide, may also act as antiepileptogenic agents through effect on glutamate neurotransmission. Such agents would be appropriate compounds for further mechanistic evaluation in animals and clinical studies in PTE patients if the studies proposed here suggest that chronic dysregulation of glutamatergic release is associated with epileptogenesis. Pretransfection with a recently developed adeno-associated virus for the GLT-1 transporter to enhance glutamate reuptake, currently under investigation by one of us, may provide yet another, novel approach to target a specific mechanism involved in glutamate regulation should these studies confirm a glutamate transporter abnormality is associated with development of PTE. PUBLIC HEALTH RELEVANCE: In addition to affecting civilian populations, traumatic brain injury (TBI) has been labeled the signature injury of the current combat in Iraq and Afghanistan. TBI is a leading cause of morbidity and mortality in the United States and is a risk factor for post-traumatic epilepsy (PTE). Examining TBI-induced dysregulation of glutamate neurotransmitter release and reuptake, the specific aims of this proposal, may reveal important mechanisms, identify potential therapeutic targets, and pinpoint the therapeutic window to eliminate or reduce the probability of developing PTE. Recognition of molecular targets for new therapies that would extinguish TBI- induced epileptogenesis would alleviate the human suffering of civilians, soldiers and veterans as they return to civilian life, as well as diminish the concomitant long-term economic costs of epilepsy to the US health care system, the military and the Veterans Administration.